The surfaces of most mammalian cells are rich in glycoconjugates and many pathogens have exploited the terminal carbohydrate of these glycoconjugates for cell attachment during the initial stages of pathogenesis. By way of example, the viral pathogens influenza and parainfluenza both bind to sialic acid receptors at the mammalian cell surface.
Sialic acid recognition is mediated via lectins or lectin-like molecules and their corresponding receptors (3). In most cases, sialic acid binding lectins, which also include several viral glycoproteins and bacterial toxins (2,4), as well as the mammalian lectin superfamilies such as the siglecs (5) and selectins (6), bind to their receptor with relatively high affinity due to the multivalent nature of these molecules, thus alleviating the low intrinsic affinity that most protein-carbohydrate interactions are associated with (7,8). Generally, association constants (Ka) for the binding of monovalent and divalent sialosides by such lectins can reach 104M−1. However, by virtue of their multivalency, some sialic acid binding lectins can interact with multivalent cell surface glycans to achieve affinities reaching 109M−1 by an avidity effect. These enhanced affinities have been shown in part to be due to improved structural packing of proteins promoted by ligand binding, associated with favourable binding energetics (9-11). One of the best-studied multivalent lectin-sialic acid interactions is the influenza virus trimeric hemagglutinin, which can achieve affinities up to 108M−1 compared to around 4×102M−1 when one, or both of the entities are not in a multivalent state (12).
Sialidases, or neuraminidases, catalyze the hydrolysis of sialic acids from a variety of glycoconjugates and are often modular enzymes, containing accessory modules attached to the catalytic core of the protein. Some of these modules have been identified as carbohydrate binding modules (CBMs). CBMs are found widely in glycoside hydrolases and are discrete, non-catalytic modules that primarily exist to target the parent enzyme to its substrate for efficient hydrolysis by increasing the concentration of the enzyme at the substrate surface (13). The modules can be single, tandem or in multiples within the glycosyl hydrolase architecture. Studies have shown that they can bind to their specific glycans independently when isolated from the parent molecule, and can behave in a cooperative manner when isolated in tandem (14,15). Currently, CBMs are grouped into 52 families based upon primary sequence similarity (http://www.cazy.org/fam/acc_CBM.html). Subtle differences in the structures of CBMs can lead to diverse ligand specificity, which make CBMs an attractive system for eludicating protein-carbohydrate mechanisms.
A poster entitled “Engineering Multivalent Sialic Acid Recognition using the CBM40 module from Vibrio cholerae Sialidase” (published 17 May 2008: see http://www.biochem.emory.edu/conferences/glycot/Images/GlycoTProgram-Posters.pdf) describes the development of reagents with increased affinity for sialic acid through multivalency. However, the poster does not disclose that such reagents have any application in the treatment of diseases and/or conditions caused by pathogens.
It is well documented that there is increasing resistance to currently available influenza antivirals (in particular, Roche's Tamiflu) and this has emphasized the need to look at alternative methods to combat influenza. Previous studies have indicated the use of non-toxic lectins, such as SNA lectin from the elderberry as an influenza virus inhibitor but this demonstrated weak binding to sialic acid and required the presence of two or three different sugar moieties for recognition and effective inhibition. Recent work using a recombinant sialidase-fusion protein designated DAS181 (Fludase, developed by NexBio Inc.) is currently being investigated as another of these alternatives. This protein effectively removes sialic acids from the surface of epithelial cells, rendering the virus unable to bind to receptors. However, by removing sialic acids using a sialidase, this can also expose cryptic receptors, which may serve as receptors for other pathogens.
The present invention aims to provide compounds, compositions, medicaments and methods useful in the treatment of diseases and/or conditions caused by pathogens and to obviate the problems associated with the prior art.